Sweet gene
The confirmation experiment of Sweet gene.
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The confirmation experiment of Sweet gene.pdf
We get the Sweet gene groE which could help E-coli folding the protein correctly at high temperature from the genome of BL21. Caring this protein would make the host bacteria grow better at high temperature.
![](https://static.igem.org/mediawiki/2012/8/8d/Sweet_gene1.jpg)
Experimental procedure:
Step 1: recover groE and Blank ( E.coli without sweet gene),and shake them at 37℃ for 12 to 16h at 150rpm.
Step 2: measure the OD600 of bacterial liquid .
Step 3: use 50ul bacterial liquid which has been diluted times to coat plates. Set temperature and induced factor (IPTG, 1 mM) control.
Step 4: add 5ul bacterial liquid which has been diluted times to liquid Luria-Bertani medium .Set temperature control. After cultivating for 12 to 16h, measure the OD600 of bacterial liquid.
![](https://static.igem.org/mediawiki/2012/thumb/f/f9/Sweet_gene_Fig.1.jpg/800px-Sweet_gene_Fig.1.jpg)
Fig1. The experimental procedure of Sweet gene’s confirmatory experiment
The data of Sweet gene’s confirmation experiment version 1.0
![](https://static.igem.org/mediawiki/2012/thumb/b/bb/Sweet_gene.Fig2.jpg/800px-Sweet_gene.Fig2.jpg)
Fig2. The initial OD600 of the groE and Blank
![](https://static.igem.org/mediawiki/2012/thumb/2/2b/Sweet_gene.Fig3.jpg/800px-Sweet_gene.Fig3.jpg)
Fig3. The OD600 of the E.coli after 16h
![](https://static.igem.org/mediawiki/2012/thumb/9/9a/Sweet_gene.Fig4.jpg/800px-Sweet_gene.Fig4.jpg)
Fig4. The average value of the OD600 of fig 2
The data of Sweet gene’s confirmation experiment version 2.0
The initial OD600 of the g+p ( groE on the backbone of pET32a) and p( plasmid pET32a)
![](https://static.igem.org/mediawiki/2012/c/ca/Seua_Sweet_gene.5.jpg)
After 4 h, we measure the OD600 of each kind of cell liquid:
![](https://static.igem.org/mediawiki/2012/thumb/8/83/Sweet_gene.6.jpg/600px-Sweet_gene.6.jpg)
Dealing with these data, we got the results as follows:
![](https://static.igem.org/mediawiki/2012/thumb/b/bd/Sweet_gene.7.jpg/569px-Sweet_gene.7.jpg)
Averaging the different volume, we have a more visual figure. The Sweet gene plays a powerful role in the temperature struggle.
![](https://static.igem.org/mediawiki/2012/7/77/Sweet_gene.8.jpg)
After 17 hours:
![](https://static.igem.org/mediawiki/2012/thumb/c/c0/Sweet_gene.9.jpg/600px-Sweet_gene.9.jpg)
The average of data after 17h:
![](https://static.igem.org/mediawiki/2012/thumb/b/bc/Sweet_gene.10.jpg/504px-Sweet_gene.10.jpg)
After 17h, we found a strange phenomenon. The g+p lives better at 42℃ without IPTG, and we don’t know the reason. Besides, the Sweet gene still works in a temperature struggle.
![](https://static.igem.org/mediawiki/2012/4/43/Sweet_gene.11.jpg)
After 45h:
![](https://static.igem.org/mediawiki/2012/thumb/b/ba/Sweet_gene.12.jpg/605px-Sweet_gene.12.jpg)
The average of data after 45h:
![](https://static.igem.org/mediawiki/2012/thumb/d/d4/Sweet_gene.13.jpg/542px-Sweet_gene.13.jpg)
After 45h, the strange phenomenon became stranger. There was no regularity anymore. The Sweet gene was no longer performing its function.
![](https://static.igem.org/mediawiki/2012/b/b0/Sweet_gene.14.jpg)
We still compared the OD600 of one type of bacteria while time elapses.
The volume of bacterial liquid is 5 ul.
![](https://static.igem.org/mediawiki/2012/3/34/Sweet_gene.15.jpg)
The volume of bacterial liquid is 10 ul.
![](https://static.igem.org/mediawiki/2012/e/e8/Sweet_gene.16.jpg)
The volume of bacterial liquid is 15 ul.
![](https://static.igem.org/mediawiki/2012/d/db/Sweet_gene.17.jpg)
From these figures, we still got the same result as the data above.
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